Abstract
The eukaryotic DNA replication fork is a hub of enzymes that continuously act to synthesize DNA, propagate DNA methylation and other epigenetic marks, perform quality control, repair nascent DNA, and package this DNA into chromatin. Many of the enzymes involved in these spatiotemporally correlated processes perform their functions by binding to proliferating cell nuclear antigen (PCNA). A long-standing question has been how the plethora of PCNA-binding enzymes exert their activities without interfering with each other. As a first step towards deciphering this complex regulation, we studied how Chromatin Assembly Factor 1 (CAF-1) binds to PCNA. We demonstrate that CAF-1 binds to PCNA in a heretofore uncharacterized manner that depends upon a cation-pi (π) interaction. An arginine residue, conserved among CAF-1 homologs but absent from other PCNA-binding proteins, inserts into the hydrophobic pocket normally occupied by proteins that contain canonical PCNA interaction peptides (PIPs). Mutation of this arginine disrupts the ability of CAF-1 to bind PCNA and to assemble chromatin. The PIP of the CAF-1 p150 subunit resides at the extreme C-terminus of an apparent long α-helix (119 amino acids) that has been reported to bind DNA. The length of that helix and the presence of a PIP at the C-terminus are evolutionarily conserved among numerous species, ranging from yeast to humans. This arrangement of a very long DNA-binding coiled-coil that terminates in PIPs may serve to coordinate DNA and PCNA binding by CAF-1.